Simultaneous determination of doxifluridine and 5-fluorouracil in monkey serum by high performance liquid chromatography with tandem mass spectrometry

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Analytical technologies in the biomedical and life sciences, 2008-11, Vol.875 (2), p.487-492
• Main Authors: Woo, Young-Ah, Kim, Ghee-Hwan, Jeong, Eun Ju, Kim, Choong-Yong
• Format: Article
• Language: English
• Subjects: 5-FU; Administration, Oral; Analysis; Analytical, structural and metabolic biochemistry; Animals; Antimetabolites, Antineoplastic - blood; Antimetabolites, Antineoplastic - pharmacokinetics; Biological and medical sciences; Chromatography, High Pressure Liquid - methods; Doxifluridine; Drug Stability; Floxuridine - blood; Floxuridine - pharmacokinetics; Fluorouracil - blood; Fundamental and applied biological sciences. Psychology; General pharmacology; LC/MS/MS; Linear Models; Macaca fascicularis; Male; Medical sciences; Monkey; Pharmacology. Drug treatments; Reproducibility of Results; Sensitivity and Specificity; Serum; Spectrometry, Mass, Electrospray Ionization - methods; Tandem Mass Spectrometry - methods
• ISSN: 1570-0232; 1873-376X
• DOI: 10.1016/j.jchromb.2008.09.037

A reverse-phase high performance liquid chromatography method with electrospray ionization and detection by mass spectrometry is described for the simultaneous determination of doxifluridine and its active metabolite 5-fluorouracil in monkey serum. A liquid/liquid extraction with ethyl acetate (90%) and isopropyl alcohol (10%) was used to extract simultaneously doxifluridine and 5-FU which have considerable difference in the polarity. Optimum chromatographic separation was achieved on a Agilent Zorbax C 18 (100 mm × 2.1 mm, 3.5 μm) column with a mobile phase of methanol–water (20:80, v/v). The flow rate was 0.2 mL/min with total cycle time of 5 min. The lower limit of quantification (LLOQ) was validated at 10.0 ng/mL of serum for both doxifluridine and 5-FU. Accuracy and precision of quality control (QC) samples for both compounds met FDA Guidance criteria of ±15% with average QC accuracy of 95.5–105.0% and coefficients of variation of 1.1–9.5% in the 10–2000 ng/mL concentration range. This method demonstrated adequate sensitivity, specificity, accuracy, precision, stability to support the analysis of monkey serum samples.